Dataset Information

New Gene Markers for Metabolic Processes and Homeostasis in Porcine Buccal Pouch Mucosa during Cells Long Term-Cultivation-A Primary Culture Approach.

ABSTRACT: The oral mucosal tissue is a compound structure composed of morphologically and physiologically different cell types. The morphological modification involves genetically determined lifespan, which may be recognized as the balance between cell survival and apoptosis. Although the biochemical processes and pathways in oral mucosa, with special regards to drug transport, delivery, and metabolism, are well known, the cellular physiological homeostasis in this tissue requires further investigation. The porcine buccal pouch mucosal cells (BPMCs) collected from 20 pubertal crossbred Landrace gilts, were used in this study. Immediately after recovery, the oral mucosa was separated micro-surgically, and treated enzymatically. The dispersed cells were transferred into primary in vitro culture systems for a long-term cultivation of 30 days. After each step of in vitro culture (IVC), the cells were collected for isolation of total RNA at 24 h, 7, 15, and 30 days of IVC. While the expression was analyzed for days 7, 15, and 30, the 24th hour was used as a reference for outcome calibration. The gene expression profile was determined using Affymetrix microarray assays and necessary procedures. In results, we observed significant up-regulation of SCARB1, PTGS2, DUSP5, ITGB3, PLK2, CCL2, TGFB1, CCL8, RFC4, LYN, ETS1, REL, LIF, SPP1, and FGER1G genes, belonging to two ontological groups, namely "positive regulation of metabolic process", and "regulation of homeostatic process" at 7 day of IVC as compared to down-regulation at days 15 and 30. These findings suggest that the metabolic processes and homeostatic regulations are much more intense in porcine mucosal cells at day 7 of IVC. Moreover, the increased expression of marker genes, for both of these ontological groups, may suggest the existence of not only "morphological lifespan" during tissue keratinization, but also "physiological checkpoint" dedicated to metabolic processes in oral mucosa. This knowledge may be useful for preclinical experiments with drugs delivery and metabolism in both animals and humans.

SUBMITTER: Dyszkiewicz-Konwinska M

PROVIDER: S-EPMC5979461 | biostudies-literature | 2018 Mar

REPOSITORIES: biostudies-literature

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New Gene Markers for Metabolic Processes and Homeostasis in Porcine Buccal Pouch Mucosa during Cells Long Term-Cultivation-A Primary Culture Approach.

Dyszkiewicz-Konwińska Marta M Nawrocki Mariusz J MJ Huang Yan Y Bryja Artur A Celichowski Piotr P Jankowski Maurycy M Błochowiak Katarzyna K Mehr Katarzyna K Bruska Małgorzata M Nowicki Michał M Zabel Maciej M Kempisty Bartosz B

International journal of molecular sciences 20180329 4

The oral mucosal tissue is a compound structure composed of morphologically and physiologically different cell types. The morphological modification involves genetically determined lifespan, which may be recognized as the balance between cell survival and apoptosis. Although the biochemical processes and pathways in oral mucosa, with special regards to drug transport, delivery, and metabolism, are well known, the cellular physiological homeostasis in this tissue requires further investigation. T ...[more]

PMID: 29596348

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Project description:BackgroundGene expression changes resulting from conditions such as disease, environmental stimuli, and drug use, can be monitored in the blood. However, a less invasive method of sample collection is of interest because of the discomfort and specialized personnel necessary for blood sampling especially if multiple samples are being collected. Buccal mucosa cells are easily collected and may be an alternative sample material for biomarker testing. A limited number of studies, primarily in the smoker/oral cancer literature, address this tissue's efficacy as an RNA source for expression analysis. The current study was undertaken to determine if total RNA isolated from buccal mucosa could be used as an alternative tissue source to assay relative gene expression.MethodsTotal RNA was isolated from swabs, reverse transcribed and amplified. The amplified cDNA was used in RT-qPCR and microarray analyses to evaluate gene expression in buccal cells. Initially, RT-qPCR was used to assess relative transcript levels of four genes from whole blood and buccal cells collected from the same seven individuals, concurrently. Second, buccal cell RNA was used for microarray-based differential gene expression studies by comparing gene expression between a group of female smokers and nonsmokers.ResultsAn amplification protocol allowed use of less buccal cell total RNA (50 ng) than had been reported previously with human microarrays. Total RNA isolated from buccal cells was degraded but was of sufficient quality to be used with RT-qPCR to detect expression of specific genes. We report here the finding of a small number of statistically significant differentially expressed genes between smokers and nonsmokers, using buccal cells as starting material. Gene Set Enrichment Analysis confirmed that these genes had a similar expression pattern to results from another study.ConclusionsOur results suggest that despite a high degree of degradation, RNA from buccal cells from cheek mucosa could be used to detect differential gene expression between smokers and nonsmokers. However the RNA degradation, increase in sample variability and microarray failure rate show that buccal samples should be used with caution as source material in expression studies.

Project description:BackgroundFemale urethral stricture (FUS) represents a sporadic condition. There is a lack of data and standardized guidelines on diagnostics and therapeutics. Several surgical techniques have been described for FUS urethroplasty, among which the flap-based or graft-based ones are most reported. Buccal mucosa graft (BMG) represents the gold standard for male urethroplasty, and this can theoretically be applied also to FUS treatment.ObjectiveTo describe and present preliminary results of a novel minimally invasive technique for buccal mucosa dorsal graft (mini-dorsal BMG) urethroplasty for the treatment of FUS.Design setting and participantsThis is a retrospective study on buccal mucosa dorsal graft urethroplasty for the treatment of FUS.Surgical procedureEvery patient was placed in lithotomic position. Two stiches were placed at 10 and 2 o'clock positions to facilitate the dorsal median urethrotomy. The margins of the incised dorsal urethra at the 12 o'clock position are then dissected from the periurethral tissue. This dissection results in an elliptical raw area between the edges of the urethra over the periurethral tissue. The harvested BMG was fixed with several quilting sutures, using 5-0 and 4-0 absorbable sutures, to cover the raw area. The margins of the graft were sutured to the edges of the incised urethra.MeasurementsA chart review was performed.Results and limitationsThirteen patients underwent the mini-dorsal-BMG technique. The median preoperative uroflow was 5.6 (3-13) ml/s, and the median postoperative value was 23.4 (14-58) ml/s.ConclusionsThe mini-dorsal-BMG technique for the treatment of FUS gives good results with low complication rates. Other series and long-term follow-up are necessary to confirm the reproducibility of this technique.Patient summaryWe present the technical aspects and the promising preliminary results of a novel surgical technique for the treatment of female urethral stricture by using the buccal mucosa to correct this invalidating disease.

Dataset Information

New Gene Markers for Metabolic Processes and Homeostasis in Porcine Buccal Pouch Mucosa during Cells Long Term-Cultivation-A Primary Culture Approach.

Publications

New Gene Markers for Metabolic Processes and Homeostasis in Porcine Buccal Pouch Mucosa during Cells Long Term-Cultivation-A Primary Culture Approach.

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